Cells were isolated from the major arteries of 17-day chick embryos by digestion of the tissue with collagenase and trypsin. The cells, when examined immediately after isolation, exhibited a high degree of viability and they were shown to synthesize and secrete procollagen at a high and constant rate for several hours when incubated in suspension in modified Krebs medium. Continuous labelling of the cells with [14C]proline demonstrated a lag of about 30min between the time at which the synthesis of non-diffusible peptide-bound hydroxy[14C]proline became linear and the time at which its secretion into the medium became linear. This lag time compares with that of 18min observed for freshly isolated matrix-free cells from embryonic-chick tendon, which synthesize and secrete the same type of collagen. Gel-filtration chromatography and polyacrylamide-gel electrophoresis indicated that the collagenous polypeptides secreted into the medium were in the precursor form, known as procollagen, and that the constituent pro-α-chains were linked by interchain disulphide bonds and were also in a triple-helical conformation. Characterization of the secreted procollagen by gel-filtration chromatography, polyacrylamide-gel electrophoresis, DEAE-agarose chromatography, and polyacrylamide-gel electrophoresis of peptides obtained by CNBr cleavage, indicated that the predominant form was type-I procollagen. This work extends the range of freshly isolated matrix-free cell systems, which have been characterized for use in studies on the biosynthesis and secretion of procollagen, and it indicates differences in the rates of secretion of procollagen in different cell types secreting the same type of procollagen.
Research Article|September 15 1977
Synthesis of procollagen by matrix-free cells from embryonic-chick arteries
J. David Schofield;
J. David Schofield
*Department of Medical Biochemistry, University of Manchester Medical School, Manchester M13 9PT, U.K.
†Geigy Unit for Research into Aging, Department of Geriatric Medicine, University Hospital of South Manchester, Manchester M20 8LR, U.K.
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Biochem J (1977) 166 (3): 437-446.
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J. David Schofield, Richard Harwood, David S. Jackson; Synthesis of procollagen by matrix-free cells from embryonic-chick arteries. Biochem J 15 September 1977; 166 (3): 437–446. doi: https://doi.org/10.1042/bj1660437
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